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BACKGROUND The CLL11 trial evaluated front line treatment with Clb vs. R-Clb vs. G-Clb in 781 physically unfit patients. Genomic aberrations, IGHV and TP53 mutation status are established prognostic factors in CLL, while NOTCH1, SF3B1, ATM and othersare recurrently mutated genes of potential prognostic and/or predictive value. METHODS We performed amplicon-based targeted next generation sequencing of all coding exons (TP53, ATM, MYD88, FBXW7, BIRC3, XPO1, POT1) or hotspots (NOTCH1 exons 33/34, SF3B1 exons 14-16/18) via Illumina TSCA™ in a representative subset of 689 (88.2%) patients of the CLL11 trial at baseline. Mean read depth was 1363x (range 636-1879) and a variant allele frequency >5% was determined as mutation. RESULTS Incidences of gene mutations were: ATM 19.6%, NOTCH1 18.6%, SF3B1 12.9%, TP53 10.7%, POT1 7.3%, XPO1 6.8%, BIRC3 5.2%, FBXW7 3.8%, and MYD88 3.3%. We found several significant associations with clinical and genetic baseline characteristics including most prominently IGHVunmut with ATMmut (p=0.013), NOTCH1mut (p<0.001), POT1mut (p=0.026), XPO1mut (p<0.001), and MYD88wt (p<0.001). ATMmut and NOTCH1mut were more frequent in Binet stage B/C (p=0.011, p=0.001, resp.). POT1mut showed significantly higher and FBXW7mut lower WBC (p=0.033, p=0.002, resp.). Finally, we found associations of ATMmut with del11q, TP53mut with del17p, NOTCH1mut and FBXW7mut with +12q (p<0.001, each), BIRC3mut and POT1wt with +12q (p=0.003, p=0.014, resp.). Regarding response to treatment, TP53mut was associated with reduced ORR for R-Clb (p<0.001) and G-Clb (p=0.002), BIRC3mut for R-Clb (p=0.001) and FBXW7mut for G-Clb (p=0.045) only. MRD negativity (<10-4) at the end of treatment was more frequently achieved by G-Clb as compared with R-Clb in the subgroups ATMmut (p<0.001), BIRC3mut (p=0.007), NOTCH1mut (p<0.001) and FBXW7mut (p=0.047). At a median observation time of 41.9 months, there were 529 (76.8%) events for PFS and 195 (28.3%) events for OS in the cohort studied. G-Clb prolonged PFS as compared to R-Clb in the overall cohort (median 27.1 vs. 15.7 months, HR 0.49, p<0.001) and in all gene mutation subgroups, except for FBXW7mut. Subgroups defined by ATMmut and TP53mut (Fig. 1) had significantly reduced PFS in both R-Clb and G-Clb (ATMmut HR 1.41, p=0.029/HR 1.67, p=0.004; TP53mut HR 3.36/HR 2.28, p<0.001, each). For SF3B1mut and XPO1mut a non-significant trend was observed for shorter PFS in both treatment arms. Notably, NOTCH1mut (Fig. 2)was associated with decreased PFS only in the R-Clb arm (HR 1.42, p=0.03), but not in Clb monotherapy (HR 1.52, p=0.103) or G-Clb arm (HR 1.08, p=0.697). Similarly, POT1mut (HR 1.69, p=0.043) and BIRC3mut (HR 1.69, p=0.023) affected PFS only in the R-Clb arm, and FBXW7mut exclusivelyin the G-Clb arm (HR 2.35, p=0.028). We performed multivariable Cox regression analysis including clinical and genetic prognostic parameters that were significant in univariate analyses to evaluate their independent prognostic value for PFS and OS. For PFS we identified G-Clb vs. Clb (HR 0.21, p<0.001), G-Clb vs. R-Clb (HR 0.42, p<0.001), 17p- (HR 1.57, p=0.038), IGHVunmut (HR 2.19, p<0.001), TP53mut (HR 2.19, p<0.001), ATMmut (HR 1.33, p=0.009), and serum thymidine kinase (TK) >10 U/l (HR 1.22, p=0.037) as independent prognostic factors. Regarding OS, 17p- (HR 2.57, p<0.001), 11q- (HR 1.58, p=0.015), IGHVunmut (HR 2.0, p<0.001), POT1mut (HR 2.17, p=0.001), age >80 years (HR 1.68, p=0.006), Binet stage C (HR 2.1, p<0.001), ECOG ≥1 (HR 1.6, p=0.007), and TK >10 U/l (HR 1.59, p=0.004) were identified as independent adverse prognostic factors, while G-Clb vs. Clb (HR 0.61, p=0.022) and G-Clb vs. R-Clb (HR 0.69, p=0.029) correlate with a better OS. CONCLUSION The advent of targeted next generation sequencing has led to a more comprehensive molecular characterization of CLL patients and can provide the basis for genotype specific treatment concepts. G-Clb improves treatment outcome as compared to R-Clb for most subgroups defined by gene mutations and overcomes NOTCH1mut-associated Rituximab resistance. In particular, addition of G achieved more MRD negativity in NOTCH1mut, ATMmut, BIRC3mut and FBXW7mut patients. Both ATMmut and TP53mut remain strong prognostic factors. New treatment regimens with novel agents should be considered for patients with adverse independent prognostic factors for PFS and OS, most notably 17p-, 11q-, IGHVunmut and TP53mut. Figure 1 Figure 1. Disclosures Tausch: Gilead: Other: Travel support, Speakers Bureau; Amgen: Other: Travel support; Celgene: Other: Travel support. Bahlo:F. Hoffman-La Roche: Honoraria, Other: Travel grant. Goede:Glaxo Smith Kline: Consultancy, Honoraria; F- Hoffmann-LaRoche: Consultancy, Honoraria, Other: Travel grants; Janssen: Consultancy, Other: Travel grants; Gilead: Consultancy; Mundipharma: Consultancy, Honoraria; Bristol Myer Squibb: Honoraria. Ritgen:Roche: Membership on an entity's Board of Directors or advisory committees, Research Funding. Langerak:InVivoScribe Technologies: Patents & Royalties: Royalties are provided to European Network (EuroClonality); F. Hofmann-LaRoche, Genentech: Research Funding. Fingerle-Rowson:F. Hoffmann-LaRoche: Employment. Kneba:Amgen: Research Funding; Gilead: Consultancy, Honoraria, Other: Travel grants, Research Funding; AbbVie: Consultancy, Honoraria, Other: Travel grants; Janssen-Cilag: Consultancy, Honoraria, Other: Travel grants; Glaxo-SmithKline: Other: Travel grants; Roche: Consultancy, Honoraria, Other: Travel grants, Research Funding. Fischer:Roche: Other: travel grants. Hallek:F. Hoffmann-LaRoche: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; AbbVie: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Mundipharma: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Janssen-Cilag: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Gilead: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau. Stilgenbauer:Janssen: Consultancy, Honoraria, Other: Travel grants , Research Funding; GSK: Consultancy, Honoraria, Other: Travel grants , Research Funding; Celgene: Consultancy, Honoraria, Other: Travel grants , Research Funding; Amgen: Consultancy, Honoraria, Other: Travel grants, Research Funding; Gilead: Consultancy, Honoraria, Other: Travel grants , Research Funding; Boehringer Ingelheim: Consultancy, Honoraria, Other: Travel grants , Research Funding; Mundipharma: Consultancy, Honoraria, Other: Travel grants , Research Funding; Hoffmann-La Roche: Consultancy, Honoraria, Other: Travel grants , Research Funding; AbbVie: Consultancy, Honoraria, Other: Travel grants, Research Funding; Pharmacyclics: Consultancy, Honoraria, Other: Travel grants , Research Funding; Genentech: Consultancy, Honoraria, Other: Travel grants , Research Funding; Genzyme: Consultancy, Honoraria, Other: Travel grants , Research Funding; Novartis: Consultancy, Honoraria, Other: Travel grants , Research Funding; Sanofi: Consultancy, Honoraria, Other: Travel grants , Research Funding.

Somatic hypermutation (SHM) is a pivotal process in adaptive immunity that occurs in the germinal centre and allows B cells to change their primary DNA sequence and diversify their antigen receptors. Here, we report that genome binding of Lamin B1, a component of the nuclear envelope involved in epigenetic chromatin regulation, is reduced during B-cell activation and formation of lymphoid germinal centres. Chromatin immunoprecipitation-Seq analysis showed that kappa and heavy variable immunoglobulin domains were released from the Lamin B1 suppressive environment when SHM was induced in B cells. RNA interference-mediated reduction of Lamin B1 resulted in spontaneous SHM as well as kappa-light chain aberrant surface expression. Finally, Lamin B1 expression level correlated with progression-free and overall survival in chronic lymphocytic leukaemia, and was strongly involved in the transformation of follicular lymphoma. In summary, here we report that Lamin B1 is a negative epigenetic regulator of SHM in normal B-cells and a ‘mutational gatekeeper’, suppressing the aberrant mutations that drive lymphoid malignancy.

Introduction: In chronic lymphocytic leukemia (CLL), improved anti-CD20 antibodies such as obinutuzumab (GA101) and targeted drugs, such as ibrutinib, show very good efficacy and tolerability. With the CLL2-BIG trial, the German CLL Study Group designed a regimen composed of obinutuzumab and ibrutinib for induction and maintenance therapy. Patients with high tumor burden receive bendamustine as debulking resulting in the proposed "sequential triple-T" concept [Hallek M., Blood 2013] of a tailored and targeted treatment aiming at total eradication of minimal residual disease (MRD). Here we present the results of the primary endpoint analysis of the trial. Methods : This prospective, open-label, multicenter phase-II trial investigates the efficacy and safety of a novel regimen for physically fit and unfit CLL patients (pts) requiring treatment, irrespective of high-risk genetics. 62 pts were to be recruited according to a predefined allocation for the two strata of first-line and relapse/refractory treatment. Six cycles of induction treatment with obinutuzumab and ibrutinib were administered followed by maintenance therapy with continuous ibrutinib and obinutuzumab every three months until achievement of an MRD-negative complete remission or up to 24 months. Pts with an absolute lymphocyte count ≥ 25.000/µl and/or lymph nodes ≥ 5cm received two cycles of bendamustine before start of induction. The primary endpoint is overall response rate (ORR) three month after the start of last induction cycle administered; secondary endpoints include ORR after debulking, MRD evaluations and safety parameters (graded per the NCI CTCAE v.4 criteria). Results: 66 pts were enrolled between January and August 2015. Five pts completed less than two cycles of induction and were therefore excluded from the full analysis set as defined by protocol. The current analysis includes 58 pts; an update including all 61 pts will be available for presentation at the meeting. The median age was 65.5 (range 36-83) years and the median time since initial diagnosis was 56.2 (2.1 - 222.8) months. 27 pts were treatment-naïve and 31 relapsed/refractory with a median of one prior treatment line (1-5). Baseline characteristics are summarized in table 1. 41 pts (71%) received bendamustine debulking, of these 27 pts (66%) responded; five pts (12%) achieved clinical complete remission (CR), 3 pts (7%) clinical CR with incomplete recovery of the bone marrow (CRi) and 19 pts (46%) partial remission (PR). 57 pts completed six cycles of induction treatment. One patient died after the fifth course due to grade 5 duodenitis related to study therapy. The combination showed promising efficacy with an ORR of 100% by investigator assessment at the end of induction. Statistically, the primary endpoint was met and the null hypothesis could be rejected. Response rates are presented in table 2. 27 pts (47%) achieved MRD negativity (< 10-4 by flow cytometry) in peripheral blood (pB) at the end of induction. An intermediate MRD-status (≥ 10-4 and < 10-2) was found in 15 pts (26%) whereas 13 pts (22%) were MRD positive (≥ 10-2). In three pts (5%) no MRD sample was available. So far, 38 pts (65%) received at least one dose of maintenance treatment; one patient already stopped treatment due to MRD negativity as defined per protocol. CTC Grade 3-4 adverse events occurred in 19 pts (33%) during induction therapy. The most common toxicities observed are shown in table 3. *The table includes all grade 3-4 AEs regardless of frequency and AEs of any grade observed in at least six patients. Conclusion: With an ORR of 100% and an MRD negativity rate of 47% in the pB the BIG-regimen showed a very good efficacy in a heterogeneous study population. No major toxicity occurred so far. Table 1. Demographics and baseline characteristics Table 1. Demographics and baseline characteristics Table 2. Response rates Table 2. Response rates Table 3 Safety Table 3. Safety Disclosures von Tresckow: Janssen-Cilag: Honoraria, Other: Travel grants, Research Funding; Celgene: Other: Travel grants; Hoffmann-LaRoche: Other: Travel grants, Research Funding. Cramer:Astellas: Other: Travel grants; Janssen-Cilag: Honoraria, Other: Travel grants, Research Funding; GlaxoSmithKline/Novartis: Research Funding; Gilead: Other: Travel grants, Research Funding; Hoffmann-LaRoche: Honoraria, Other: Travel grants, Research Funding; Mundipharma: Other: Travel grants. Bahlo:F. Hoffman-La Roche: Honoraria, Other: Travel grant. Engelke:Hoffmann-LaRoche: Other: Travel grants. Langerbeins:Hoffmann-LaRoche: Honoraria, Other: Travel grants, Research Funding; Janssen-Cilag: Honoraria, Other: Travel grants, Research Funding; Mundipharma: Honoraria, Other: Travel grants, Research Funding. Fink:Mundipharma: Other: Travel grants; Celgene: Other: Travel grants, Research Funding; AbbVie: Other: Travel grants; Hoffmann-LaRoche: Other: Travel grants. Illmer:Hoffmann-LaRoche: Honoraria, Other: travel grants. Ritgen:Roche: Membership on an entity's Board of Directors or advisory committees, Research Funding. Fischer:Hoffmann-LaRoche: Other: Travel grants. Wendtner:Mundipharma: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; Janssen-Cilag: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; GlaxoSmithKline: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; Hoffmann-LaRoche: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Travel grants, Research Funding. Stilgenbauer:Janssen: Consultancy, Honoraria, Other: Travel grants , Research Funding; Hoffmann-La Roche: Consultancy, Honoraria, Other: Travel grants , Research Funding; Pharmacyclics: Consultancy, Honoraria, Other: Travel grants , Research Funding; Amgen: Consultancy, Honoraria, Other: Travel grants, Research Funding; GSK: Consultancy, Honoraria, Other: Travel grants , Research Funding; Mundipharma: Consultancy, Honoraria, Other: Travel grants , Research Funding; Genentech: Consultancy, Honoraria, Other: Travel grants , Research Funding; Boehringer Ingelheim: Consultancy, Honoraria, Other: Travel grants , Research Funding; Celgene: Consultancy, Honoraria, Other: Travel grants , Research Funding; Novartis: Consultancy, Honoraria, Other: Travel grants , Research Funding; Sanofi: Consultancy, Honoraria, Other: Travel grants , Research Funding; Gilead: Consultancy, Honoraria, Other: Travel grants , Research Funding; AbbVie: Consultancy, Honoraria, Other: Travel grants, Research Funding; Genzyme: Consultancy, Honoraria, Other: Travel grants , Research Funding. Böttcher:AbbVie: Honoraria, Research Funding; Hoffmann-LaRoche: Honoraria, Other: Travel grants, Research Funding; Celgene: Research Funding. Eichhorst:Abbvie: Consultancy; Mundipharma: Consultancy, Research Funding, Speakers Bureau; Roche: Consultancy, Research Funding, Speakers Bureau. Hallek:Amgen: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Janssen-Cilag: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Gilead: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; Mundipharma: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; F. Hoffmann-LaRoche: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau; AbbVie: Consultancy, Honoraria, Other: travel support, Research Funding, Speakers Bureau.